MedWorm.com provides a medical RSS filtering service. Over 6000 RSS medical sources are combined and output via different filters. This feed contains the latest items from the 'BJ Disease' source. Sun, 21 Mar 2010 16:57:40 +0100 http://www.medworm.com/index.php?rid=3349672&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20100016 Previous studies showed that apolipoprotein E (apoE) expression in macrophages suppresses inflammatory response. Whether the endogenously synthesized apoE acts intracellularly or after its secretion in suppressing macrophage inflammation remains unclear. The current study used the murine macrophage cell line RAW 264.7 to examine the influence of exogenous apoE on macrophage inflammatory responses induced by toll-like receptor (TLR)-4 and TLR-3 agonists lipopolysaccharide (LPS) and poly(I-C), respectively. Results showed that exogenously added apoE suppressed LPS- and poly(I-C)-induction of interleukin (IL)-6, IL-1β, and tumor necrosis factor (TNF)-α secretion by RAW 264.7 cells. The mechanism was related to apoE suppression of TLR agonist-induced phosphorylation of c-Jun N-te... BJ Disease journals http://www.medworm.com/rss/comments.php?id=3349672 Wed, 10 Mar 2010 00:00:00 +0100 3349672 http://www.medworm.com/index.php?rid=3330726&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20091177 In conclusion, Cdx2 reduced Shh expression by binding to the unmethylated Shh promoter in the intestinal metaplastic mucosa of Cdx2-transgenic mouse stomach. (Source: BJ Disease) BJ Disease journals http://www.medworm.com/rss/comments.php?id=3330726 Thu, 04 Mar 2010 00:00:00 +0100 3330726 http://www.medworm.com/index.php?rid=3310800&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20091607 Phosphatidylglycerol is a widely used mimetic to study the effects of antimicrobial peptides (AMPs) on the bacterial cytoplasmic membrane. It turned out, however, that the antibacterial activities of novel NK-2-derived AMPs could not sufficiently explained by using this simple model system. Since the lipopolysaccharide (LPS) containing outer membrane is the first barrier of Gram-negative bacteria, here we investigated interactions of NK-2 and a shortened variant thereof with viable Escherichia coli WBB01 and Proteus mirabilis R45, and with model membranes composed of LPS isolated from these two strains. Differences in net charge and charge distribution of the two LPS have been made responsible for the differential sensitivity of the respective bacteria to other AMPs. As imaged by TEM and A... BJ Disease journals http://www.medworm.com/rss/comments.php?id=3310800 Fri, 26 Feb 2010 00:00:00 +0100 3310800 http://www.medworm.com/index.php?rid=3275322&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20100074 There is a need to develop inhibitors of mosquito-borne flaviviruses, including West Nile virus (WNV). Here, we describe a novel and efficient recombinant antibody technology that led us to the isolation of the inhibitory, high affinity, human antibodies to the active site region of a viral proteinase. As a proof-of-principal, we have successfully used this technology and the synthetic naive human antibody library HuCAL GOLD to isolate selective and potent function-blocking, active site-targeting antibodies to the two-component WNV NS2B-NS3 serine proteinase, the only proteinase encoded by the flaviviral genome. First, we used the wild-type enzyme in the antibody screens. The positive antibody clones were next counter-screened using the NS2B-NS3 mutant with a single mutation of the catalyt... BJ Disease journals http://www.medworm.com/rss/comments.php?id=3275322 Tue, 16 Feb 2010 00:00:00 +0100 3275322 http://www.medworm.com/index.php?rid=3258182&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20100043 In conclusion, electrostatic interaction between ExoS and 14-3-3 via polar residues (S416, H418, D424 and D427) appears to be of secondary importance. Thus, the interaction between the “roof” of the groove of 14-3-3 and ExoS relies more on hydrophobic interaction forces, which probably contributes to induce cell death after ExoS infection and activation. (Source: BJ Disease) BJ Disease journals http://www.medworm.com/rss/comments.php?id=3258182 Tue, 09 Feb 2010 00:00:00 +0100 3258182 http://www.medworm.com/index.php?rid=3219775&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20091888 To assess the potential of mutations from the L1 loop of the tumour suppressor p53 as second-site suppressors, the effect of H115N and S116M on the p53 “hot spot” mutations has been investigated using the double mutant approach. The effects of these two mutants on the p53 “hot spots” in terms of thermal stability and DNA binding were evaluated. The results show that: (1) mutants H115N and S116M are thermally more stable than wild-type p53; (2) H115N but not S116M is capable of rescuing the DNA binding of one of the most frequent p53 mutants in cancer, R248Q, as shown by binding of R248Q/H115N to gadd45; (3) the double mutant R248Q/H115N is more stable than wild-type p53; (4) the effect of H115N as a second-site suppressor to restore DNA-binding activity is speci... BJ Disease journals http://www.medworm.com/rss/comments.php?id=3219775 Fri, 29 Jan 2010 00:00:00 +0100 3219775 http://www.medworm.com/index.php?rid=3188535&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20091097 In this report, we demonstrated that vimentin is a primary receptor required for IbeA+ E. coli K1-induced signaling and invasion of HBMEC, based on the following observations. First, E44 (IbeA+ E. coli K1 strain) invasion was blocked by vimentin inhibitors (Withfferin A and Acrylamide), a recombinant protein containing vimentin head domain and an antibody against the head domain, respectively. Secondly, overexpression of GFP-vimentin and GFP-vimentin head domain deletion mutant (VDM) significantly increased and decreased bacterial invasion, respectively. Thirdly, bacterial invasion was positively correlated with phosphorylation of vimentin at serine 82 by calcium calmodulin-dependent kinase II (CaM kinase II) and IbeA+ E. coli-induced phosphorylation of extracellular r... BJ Disease journals http://www.medworm.com/rss/comments.php?id=3188535 Wed, 20 Jan 2010 00:00:00 +0100 3188535 http://www.medworm.com/index.php?rid=3114430&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20091570 Down-regulation of the catalytic subunit of the mitochondrial H+-ATP synthase (β-F1-ATPase) is a hallmark of many human tumors. The expression level of β-F1-ATPase provides a marker of the prognosis of cancer patients as well as of the tumor response to chemotherapy. However, the mechanisms that participate in down-regulating its expression in human tumors remain unknown. Herein, we have studied the expression of β-F1-ATPase mRNA (β-mRNA) in breast, colon and lung adenocarcinomas and squamous carcinomas of the lung. Despite the down-regulation of the protein, tumor β-mRNA levels remained either unchanged (breast and lung adenocarcinomas) or significantly increased (colon and squamous lung carcinomas) when compared to paired normal tissues, suggesti... BJ Disease journals http://www.medworm.com/rss/comments.php?id=3114430 Wed, 23 Dec 2009 00:00:00 +0100 3114430 http://www.medworm.com/index.php?rid=3114429&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20091351 Legionnaires' disease is caused by a lethal colonization of alveolar macrophages with the Gram negative bacterium Legionella pneumophila. An L. pneumophila glucosyltransferase (LpGT or Lgt1) has recently been identified as a virulence factor, shutting down protein synthesis in the human cell by specific glucosylation of elongation factor 1A (EF1A), using an unknown mode of substrate recognition, and retaining glycosyl transfer. We have determined the crystal structure of LpGT in complex with substrates, revealing a GT-A fold with two unusual protruding domains. Through structure-guided mutagenesis of LpGT, several residues essential for binding of the UDP-glucose donor and EF1A acceptor substrates were identified, also affecting L. pneumophila virulence as demonstrated by microinjection st... BJ Disease journals http://www.medworm.com/rss/comments.php?id=3114429 Wed, 23 Dec 2009 00:00:00 +0100 3114429 http://www.medworm.com/index.php?rid=3084332&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20091612 We report that frataxin iron binding capacity is quite robust: even when five of the most conserved residues from the putative iron binding region are altered, at least 2 iron atoms per monomer can be bound, although with decreased affinity. Furthermore, we conclude that the acidic ridge is designed to favour function over stability. The negative charges have a functional role, but at the same time significantly impair frataxin’s stability. Removing five of those charges results in a thermal stabilization of ~24ºC and reduces the inherent conformational plasticity. Alterations on the conserved β-sheet residues have only a modest impact on the protein stability, highlighting the functional importance of residues 122-124. (Source: BJ Disease) BJ Disease journals http://www.medworm.com/rss/comments.php?id=3084332 Mon, 14 Dec 2009 00:00:00 +0100 3084332 http://www.medworm.com/index.php?rid=3007417&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20091413 Oncogenic Ras mutations render the protein constitutively active and promote tumourigenisis via chronic stimulation of effector pathways. In A549 lung adenocarcinoma approximately 50% of the total Ras population is constitutively active yet these cells display only weak activation of the effectors: ERK1/2 and Akt. In order to identify key negative regulators of oncogenic Ras signalling we performed a phosphatome RNAi screen in A549 cells and ranked their effects on phosphorylation of Ser473 of Akt. As expected, the tumour suppressor PTEN emerged as a leading hit – knockdown elevated Akt activation to 70% of maximal generated by acute EGF stimulation. Importantly, we identified other phosphatases with similar potencies including PTPN2 (TC-PTP) and PTPRJ (DEP-1/CD148). Potentiation of... BJ Disease journals http://www.medworm.com/rss/comments.php?id=3007417 Thu, 19 Nov 2009 00:00:00 +0100 3007417 http://www.medworm.com/index.php?rid=3007416&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20091062 The activity of the pro-inflammatory cytokine IL-1 is closely regulated in vivo via a variety of mechanisms, including both the control of IL-1 production and secretion as well as naturally occurring inhibitors of IL-1 function such as IL-1ra. IL-1ra is homologous to IL-1, and is able to bind but not activate the IL-1 receptor. IL-1ra can be produced by a variety of cell types, and its production is stimulated by inflammatory signals. Here we show that in macrophages the TLR mediated induction of IL-1ra from both its proximal and distal promoters involves the p38 and ERK1/2 MAPK cascades. In addition we show that mitogen and stress activated kinase 1 (MSK1) and 2, kinases activated by either ERK1/2 or p38 in vivo, are required for the induction of both IL-1ra mRNA and protein. MSKs regulat... BJ Disease journals http://www.medworm.com/rss/comments.php?id=3007416 Thu, 19 Nov 2009 00:00:00 +0100 3007416 http://www.medworm.com/index.php?rid=3007415&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090813 Cystic Fibrosis (CF) mostly follows a single F508 deletion in CFTR (ΔF508), thereby causing premature fragmentation of the nascent protein with concomitant alterations of diverse cellular functions. We show that CK2, the most pleiotropic protein kinase, undergoes allosteric control of its different cellular forms in the presence of short CFTR peptides encompassing the F508 deletion: these ΔF508 peptides drastically inhibit the isolated catalytic subunit (α) of the kinase and yet up-regulate the holoenzyme, composed of two catalytic and two non catalytic (β) subunits. Remarkable agreement between in silico docking and our biochemical data point to different sites for CFTR-ΔF peptide binding on isolated CK2α and on CK2β assembled into the holo... BJ Disease journals http://www.medworm.com/rss/comments.php?id=3007415 Thu, 19 Nov 2009 00:00:00 +0100 3007415 http://www.medworm.com/index.php?rid=3003172&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090896 A putative guanidine diphosphomannose pyrophophosphorylase (GDP-Man PP) gene from Trypanosoma brucei was identified in the genome and subsequently cloned, sequenced, and recombinantly expressed and shown to be a catalytically active dimer. Kinetic analysis revealed a Vmax of 0.34 μmol / min/ mg and Km’s of 67 μM and 12 μM for GTP and mannose-1-phosphate respectively. Further kinetic studies showed GDP-Man was a potent product feedback inhibitor. RNAi of the cytosolic TbGDP-Man PP showed mRNA levels were reduced to ~20% of wild type levels, causing the cells to die after 3-4 days, demonstrating TbGDP-Man PP is essential in the bloodstream form of T.brucei and thus a potential drug target. The RNAi induced parasites have a greatly reduced capability to form GDP-M... BJ Disease journals http://www.medworm.com/rss/comments.php?id=3003172 Wed, 18 Nov 2009 00:00:00 +0100 3003172 http://www.medworm.com/index.php?rid=2911765&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090045 Microsomal prostaglandin E synthase-1 (mPGES-1) is a stimulus-inducible enzyme that functions downstream of cyclooxygenase (COX)-2 in the prostaglandin E2 (PGE2)-biosynthetic pathway. Although COX-2-derived PGE2 is known to play a role in the development of various tumours, the involvement of mPGES-1 in carcinogenesis has not yet been fully understood. Here, we used Lewis lung carcinoma (LLC) cells with mPGES-1 knockdown or overexpression as well as mPGES-1-deficient mice to examine the roles of cancer cell- and host-associated mPGES-1 in the processes of tumorigenesis in vitro and in vivo. We found that siRNA silencing of mPGES-1 in LLC cells decreased PGE2 synthesis markedly, accompanied by reduced cell proliferation, attenuated Matrigel invasiveness and increased extracellular matrix ad... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2911765 Tue, 20 Oct 2009 23:00:00 +0100 2911765 http://www.medworm.com/index.php?rid=2911764&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20091119 The proliferation of the malaria-causing parasite Plasmodium falciparum within the erythrocyte is concomitant with massive phosphatidylcholine and phosphatidylethanolamine biosynthesis. Based on pharmacological and genetic data, de novo biosynthesis pathways of both phospholipids appear essential for parasite survival. The present study characterizes P. falciparum choline kinase (PfCK) and ethanolamine kinase (PfEK), which catalyse the first enzymatic steps of these essential metabolic pathways. Recombinant PfCK and PfEK were expressed as His-tagged fusion proteins from over-expressing E. coli strains, then purified to homogeneity and characterized. Using mice-polyclonal antibodies against recombinant kinases, PfCK and PfEK were shown to be localized within the parasite cytoplasm. Protein ... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2911764 Tue, 20 Oct 2009 23:00:00 +0100 2911764 http://www.medworm.com/index.php?rid=2893798&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090776 In this study we show that serotonin is a favoured substrate for myeloperoxidase because other physiological substrates for this enzyme, including chloride, did not affect its rate of oxidation. At low micromolar concentrations, serotonin enhanced hypochlorous acid production by both purified myeloperoxidase and neutrophils. At higher concentrations it almost completely blocked formation of hypochlorous acid. Serotonin was oxidized to a dimer by myeloperoxidase and hydrogen peroxide. It was also converted to tryptamine-4,5-dione, especially in the presence of superoxide. This toxic quinone was produced by stimulated neutrophils in a reaction that required myeloperoxidase. In plasma, stimulated human neutrophils oxidized serotonin to its dimer using the NADPH oxidase and myeloperoxidase. We... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2893798 Tue, 13 Oct 2009 23:00:00 +0100 2893798 http://www.medworm.com/index.php?rid=2889850&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20091213 In this study, we show that STAT3 binds the C-terminal tubulin associating region of SCLIP. In a search for a function of SCLIP we show that SCLIP was downregulated during OSM treatment in MCF-7 cells, which also stimulates epithelial morphology loss. SCLIP knockdown likewise triggered a loss of epithelial morphology which included reduced E-cadherin expression. We found that STAT3 was required to maintain SCLIP stability. Furthermore, inhibition of OSM-induced STAT3 activity preserved SCLIP expression and MCF-7 epithelial monolayers. Taken together, we propose that a STAT3/SCLIP interaction is required to preserved SCLIP stability and contributes to the maintenance of normal epithelial morphology. Disruption of STAT3/SCLIP interaction with OSM may contribute to cytokine mediated loss in c... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2889850 Tue, 13 Oct 2009 23:00:00 +0100 2889850 http://www.medworm.com/index.php?rid=2875139&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090343 The Gram-negative oral anaerobe Prevotella intermedia forms an iron(III) protoporphyrin IX pigment from haemoglobin. The microorganism expresses a 90 kDa cysteine protease, Interpain A (InpA), a homologue of Streptococcus pyogenes streptopain (SpeB). The role of InpA in haemoglobin breakdown and haem release was investigated. At pH 7.5, InpA mediated oxidation of oxyhaemoglobin to hydroxymethaemoglobin (in which the haem iron is oxidised to the Fe(III) state and which carries OH- as the sixth co-ordinate ligand) by limited proteolysis of globin chains as indicated by SDS-PAGE and MALDI-TOF analysis. Prolonged incubation at pH 7.5, did not result in further haemoglobin protein breakdown, but in the formation of a haemoglobin haemichrome (where the haem Fe atom is co-ordinated by another ami... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2875139 Thu, 08 Oct 2009 23:00:00 +0100 2875139 http://www.medworm.com/index.php?rid=2865099&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20091024 The brain-specific compound N-acetylaspartate (NAA) occurs almost exclusively in neurons, where its concentration reaches ≈ 20 mM. Its abundance is determined in patients by magnetic resonance spectroscopy to assess neuronal density and health. The molecular identity of the N-acetyltransferase that catalyses NAA synthesis has remained unknown, because this enzyme is membrane-bound and difficult to purify. Database searches indicated that, among the putative N-acetyltransferases (i.e., proteins homologous to known N-acetyltransferases, but with uncharacterized catalytic activity) encoded by the human and mouse genomes, two, NAT8L and NAT14, were almost exclusively expressed in brain. Transfection studies in HEK293T cells indicated that NAT8L, but not NAT14, catalysed the synthesis of... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2865099 Mon, 05 Oct 2009 23:00:00 +0100 2865099 http://www.medworm.com/index.php?rid=2777782&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20091035 The Leucine Rich Repeat Protein Kinase-2 (LRRK2) is mutated in a significant number of Parkinson’s disease patients, but little is known about its regulation and function. A common mutation changing Gly2019 to Ser enhances catalytic activity, suggesting small molecule inhibitors might have utility in treating Parkinson’s Disease. We utilised various approaches to explore the substrate specificity requirements of LRRK2 and elaborated a peptide substrate termed Nictide, that had 20-fold lower Km and nearly 2-fold higher Vmax than the widely deployed LRRKtide substrate. We demonstrate that LRRK2 has marked preference for phosphorylating Thr over Ser. We also observed that several Rho kinase (ROCK) inhibitors such as Y-27632 and H-1152, suppressed LRRK2 with similar potency to wh... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2777782 Tue, 08 Sep 2009 23:00:00 +0100 2777782 http://www.medworm.com/index.php?rid=2740791&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090424 Hemochromatosis is an iron overload disorder with age-dependent oxidative stress and dysfunction in a variety of tissues. Mutations in HFE are responsible for most cases with hemochromatosis. We recently demonstrated that Hfe is expressed exclusively in the basal membrane of retinal pigment epithelium (RPE). Here we used Hfe-/- mice to examine ferritin levels (an indirect readout for iron levels) and morphological changes in retina. We found increased ferritin accumulation in retina in 18-month-old but not in 2-month-old mice with considerable morphological damage compared to age-matched controls. The retinal phenotype included hypertrophy and hyperplasia of RPE. RPE cells isolated from Hfe-/- mice exhibited a hyperproliferative phenotype. We also compared the gene expression profile betwe... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2740791 Thu, 27 Aug 2009 23:00:00 +0100 2740791 http://www.medworm.com/index.php?rid=2715376&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090664 MAC (Mitochondrial Apoptosis-induced Channel) forms in the mitochondrial outer membrane and unleashes cytochrome c, which then orchestrates the execution of the cell. MAC opening is the commitment step of intrinsic apoptosis. Hence, closure of MAC may prevent apoptosis. Compounds that blocked release of fluorescein from liposomes by recombinant Bax were tested for their ability to directly close MAC and suppress apoptosis in FL5.12 cells. Low doses of these compounds (IC50 ranged from 19 to 966 nM) irreversibly closed MAC. These compounds also blocked cytochrome c release and halted the onset of apoptotic markers normally induced by IL-3 deprivation or staurosporine. Our results reveal the tight link amongst MAC activity, cytochrome c release, and apoptotic death, and indicate this mitocho... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2715376 Wed, 19 Aug 2009 23:00:00 +0100 2715376 http://www.medworm.com/index.php?rid=2666768&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090541 Spondyloepiphyseal dysplasia tarda (SEDT) is a late-onset X-linked recessive skeletal dysplasia caused by mutations in the gene SEDL coding for sedlin. Here, we investigate four missense mutations observed in SEDT and compare biochemical and cellular characteristics relative to the wild-type protein to address mechanism of disease and to gain insight into the function of the sedlin protein. In situ hybridization and immunohistochemical experiments in mouse growth plates revealed sedlin to be predominantly expressed in proliferating and hypertrophic chondrocytes. Cell culture studies showed the wild-type protein localized predominantly in the vicinity of the nucleus and the Golgi with further localization around the cytoplasm, whilst mutation resulted in mislocalization. The D47Y mutation e... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2666768 Sun, 02 Aug 2009 23:00:00 +0100 2666768 http://www.medworm.com/index.php?rid=2633698&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090390 The hypothesis that PDHK4 has potential as a target for the treatment of type 2 diabetes was tested by feeding wild type and PDHK4 knockout mice a high saturated fat diet that induces hyperglycemia, hyperinsulinemia, glucose intolerance, hepatic steatosis, and obesity. Previous studies have shown that PDHK4 deficiency lowers blood glucose by limiting the supply of three carbon gluconeogenic substrates to the liver. There is concern, however, that the increase in glucose oxidation caused by less inhibition of the pyruvate dehydrogenase complex by phosphorylation will inhibit fatty acid oxidation, promote ectopic fat accumulation, and worsen insulin sensitivity. This was examined by feeding wild type and PDHK4 knockout mice a high saturated fat diet for 8 months. Fasting blood glucose levels... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2633698 Wed, 22 Jul 2009 23:00:00 +0100 2633698 http://www.medworm.com/index.php?rid=2579694&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090817 FtsZ plays an essential role in bacterial cell division. We have used the assembly of FtsZ as a screen to find antibacterial agents with a novel mechanism of action. The effects of 81 compounds of 29 different structural scaffolds on FtsZ assembly in vitro were examined using a sedimentation assay. Out of these 81 compounds, 3-{5-[4-Oxo-2-thioxo-3-(3-trifluoromethyl-phenyl)-thiazolidin-5-ylidenemethyl]-furan-2-yl}-benzoic acid (OTBA) was found to promote FtsZ assembly in vitro. OTBA increased the assembly of FtsZ, caused bundling of FtsZ protofilaments, prevented dilution-induced disassembly of FtsZ protofilaments and decreased the GTPase activity in vitro. It bound to FtsZ with an apparent dissociation constant of 15 ± 1.5 µM. Further, OTBA inhibited the proliferation of Bac... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2579694 Mon, 06 Jul 2009 23:00:00 +0100 2579694 http://www.medworm.com/index.php?rid=2575824&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20082398 The hereditary spastic paraplegias (HSPs) are genetic conditions exhibiting distal degeneration of the longest axons of the corticospinal tract, resulting in spastic paralysis of the legs. The gene encoding spartin is mutated in Troyer syndrome, an HSP in which paralysis is accompanied by additional clinical features. There has been controversy over the subcellular distribution of spartin. We show here that at steady state endogenous spartin exists in a cytosolic pool that can be recruited to endosomes and to lipid droplets. Cytosolic endogenous spartin is mono-ubiquitinated and we demonstrate that it interacts via a PPXY motif with the ubiquitin E3 ligases AIP4 (WWP2) and AIP5 (WWP1). Surprisingly, neither the PPXY motif, AIP4 nor AIP5 are required for spartin’s ubiquitination and ... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2575824 Sun, 05 Jul 2009 23:00:00 +0100 2575824 http://www.medworm.com/index.php?rid=2569157&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090420 The pro-inflammatory chemokine CCL2 (MCP-1) is upregulated in the glomerular compartment during the early phase of LPS-induced nephritis. This upregulation also appears in cultured mesangial cells (MC) and is more pronounced in MC lacking the prostaglandin EP2 receptor or in MC treated with a prostaglandin EP4 receptor antagonist. To examine a possible feedback of EP receptor stimulation on CCL2 expression, we chose an in vitro model in MC with down regulated EP receptor expression. By selective overexpression of EP receptors in these cells, their effects on LPS-induced CCL2 expression were examined. Cells were stimulated with LPS and CCL2 gene expression was examined and compared to LPS stimulated, mock transfected COX-2+ cells. Overexpression of EP1 as well as EP3 had no effect o... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2569157 Wed, 01 Jul 2009 23:00:00 +0100 2569157 http://www.medworm.com/index.php?rid=2503404&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090278 Streptococcus pyogenes is one of the most common human pathogens and possesses diverse mechanisms to evade the human immune defence. One example of its immune evasion is the degradation of the chemokine IL-8 by ScpC, a serine proteinase that prevents the recruitment of neutrophils to an infection site. By applying the ANTIGENome technology and using human serum antibodies we identified Spy0416, annotated as ScpC, as a prominent antigen that induces protective immune responses in animals. We demonstrate here for the first time that the recombinant form of Spy0416 is capable of IL-8 degradation in vitro in a concentration and time dependent manner. Mutations in the conserved amino acid residues of the catalytic triad of Spy0416 completely abolished in vitro activity. However, the isolated pr... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2503404 Tue, 23 Jun 2009 23:00:00 +0100 2503404 http://www.medworm.com/index.php?rid=2503403&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090014 Receptor tyrosine kinases of the Eph family become tyrosine phosphorylated and initiate signaling events upon binding of their ligands, the ephrins. Eph receptors such as EphA2 and EphB4 are highly expressed but poorly tyrosine phosphorylated in many types of cancer cells, suggesting a limited interaction with ephrin ligands. Nevertheless, decreasing the expression of these receptors affects the malignant properties of cancer cells, suggesting that Eph receptors may influence cancer cells independently of ephrin stimulation. Ligand-independent activities of Eph receptors in cancer, however, have not been demonstrated. By using siRNA interference to downregulate EphB4 in MCF7 and MDA-MB-435 cancer cells, we found that EphB4 inhibits integrin-mediated cell substrate adhesion, spreading, and ... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2503403 Tue, 23 Jun 2009 23:00:00 +0100 2503403 http://www.medworm.com/index.php?rid=2503405&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090748 Primary hyperoxaluria type 1 (PH1) is a severe inborn disorder of glyoxylate metabolism, caused by a functional deficiency of the peroxisomal enzyme alanine:glyoxylate aminotransferase (AGXT), which converts glyoxylate to glycine using L-alanine as the amino-group donor. Even though pre-genomic studies indicate that other human transaminases can convert glyoxylate to glycine, in PH1 patients these enzymes are apparently unable to compensate for the lack of AGXT - perhaps due to their limited levels of expression, or to their localization in an inappropriate cell compartment, or to the scarcity of the required amino-group donor. Herein, we describe the cloning of eight human cytosolic aminotransferases, their recombinant expression as His6-tagged proteins and a comparative study on their ab... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2503405 Mon, 22 Jun 2009 23:00:00 +0100 2503405 http://www.medworm.com/index.php?rid=2467844&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090564 4-Hydroxynonenal (HNE), the major product of lipoperoxidation, easily reacts with proteins through adduct formation between its three main functional groups and lysyl, histidyl, and cysteinyl residues of proteins. HNE is considered to be an ultimate mediator of toxic effects elicited by oxidative stress. It can be detected in several patho-physiological conditions, in which it affects cellular processes by addition to functional proteins. We demonstrated by mass spectrometry and confirmed by immunoblotting experiments the formation of HNE-α-enolase adduct(s) in HL-60 human leukemic cells. α-Enolase is a multifunctional protein that acts as a glycolytic enzyme, transcription factor (c-myc binding protein, MBP-1) and plasminogen receptor. HNE did not affect α-enolase enz... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2467844 Tue, 09 Jun 2009 04:00:00 +0100 2467844 http://www.medworm.com/index.php?rid=2455845&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090276 This study shows for the first time that the reactions of HOSCN with low-molecular-mass thiol residues occur with rate constants in the range 7.3 × 103 M–1 s–1 (for N-acetyl-cysteine) at pH 7.4 to 7.7 × 106 M–1 s–1 (for 5-thio-2-nitrobenzoic acid at pH 6.0). An inverse relationship between the rate of reaction and the pKa of the thiol group was observed. The rates of reaction of HOSCN with thiol-containing proteins were also investigated for four proteins (creatine kinase, bovine serum albumin, β−lactoglobulin and crystallins). The values obtained for Cys residues on these proteins are in the range 1 – 7 × 104 M–1 s–1. These second-order rate constants indicate that HOSCN is a major mediator of thiol oxidat... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2455845 Wed, 03 Jun 2009 04:00:00 +0100 2455845 http://www.medworm.com/index.php?rid=2422659&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090530 The Slc30a8 gene encodes the islet-specific zinc transporter ZnT-8, which provides zinc for insulin-hexamer formation. Polymorphic variants in amino acid 325 of human ZnT-8 are associated with altered susceptibility to type 2 diabetes and ZnT-8 autoantibody epitope specificity changes in type 1 diabetes. To assess the physiological importance of ZnT-8, mice carrying a Slc30a8 exon 3 deletion were analyzed histologically and phenotyped for energy metabolism and pancreatic hormone secretion. No gross anatomical or behavioral changes or differences in body weight were observed between wild type and ZnT-8 -/- mice and ZnT-8 -/- mouse islets were indistinguishable from wild type in terms of their numbers, size and cellular composition. However, total zinc content was markedly reduced in ZnT-8 -... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2422659 Mon, 18 May 2009 04:00:00 +0100 2422659 http://www.medworm.com/index.php?rid=2413015&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090389 Dynasore, a small molecule inhibitor of dynamin, was used to probe the role of dynamin in the endocytosis of wild-type and mutant CFTR. Internalization of both wild-type and “temperature-corrected” ΔF508 CFTR was markedly inhibited by short exposure to dynasore, implicating dynamin as a key element in the endocytic internalization of both wild-type and mutant CFTR. The inhibitory effect of dynasore was readily reversible upon washout of dynasore from the growth media. Corr-4, a pharmacological corrector of ΔF508 CFTR biosynthesis, caused a marked increase in the cell surface expression of mutant CFTR. Co-incubation of ΔF508 CFTR expressing cells with corr-4 and dynasore caused a significantly greater level of cell surface CFTR than that observed in the pr... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2413015 Thu, 14 May 2009 04:00:00 +0100 2413015 http://www.medworm.com/index.php?rid=2363591&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090356 We examined ROS production in astrocytes and downstream effects leading to changes in signaling cascade, morphology, and membrane dynamics using menadione, a redox active compound capable of inducing intracellular ROS. NADPH oxidase mediated menadione-induced ROS production, which then stimulated phosphorylation of p38 mitogen-activated protein kinase (MAPK) and extracellular-signal regulated kinases (ERK1/2), and increased actin polymerization and cytoskeletal protrusions. We also showed that astrocyte plasma membranes became more molecularly-ordered under oxidative stress, which was abrogated by down regulating cytosolic phospholipase A2 (cPLA2) either with a pharmacological inhibitor or by RNA interference. In addition, a mild disruption of F-actin with cytochalasin D suppressed menadio... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2363591 Fri, 24 Apr 2009 04:00:00 +0100 2363591 http://www.medworm.com/index.php?rid=2346642&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081466 Numerous studies conducted in a diversity of adult tissues have reported that certain stem cells are characterized by the expression of a protein known as the ABCG2 transporter. In the adult pancreas, various multipotent progenitors have been proposed, however the ABCG2 marker has only been detected in the so-called Side Population. In the present study we aim to identify new ABCG2+ pancreatic cell populations and to explore whether they exhibit properties of progenitor cells. We isolated and expanded mitoxantrone-resistant cells from pancreata of lactating rats by drug selection. These cells were characterized and maintained in different stages of differentiation using several cocktail media plus matrigel. Differentiation was assessed by RT-PCR, immunocytochemistry, electron micr... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2346642 Mon, 20 Apr 2009 04:00:00 +0100 2346642 http://www.medworm.com/index.php?rid=2283096&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20082441 Genome mining and biochemical analyses have shown that L. major possesses two pathways for cysteine synthesis - the de novo biosynthesis pathway comprising serine acetyltransferase (SAT) and cysteine synthase (CS) and the reverse transsulfuration (RTS) pathway comprising cystathionine β-synthase (CBS) and cystathionine gamma-lyase (CGL). The L. major CS (LmjCS) is similar to the type A CSs of bacteria and catalyses the synthesis of cysteine using O-acetyserine and sulfide with Kms of 17.5 and 0.13 mM, respectively. LmjCS can use sulfide provided by the action of mercaptopyruvate sulfurtransferase (MST) on 3-mercaptopyruvate (3-MP). LmjCS forms a bi-enzyme complex with Leishmania SAT (and Arabidopsis SAT), with residues K222, H226 and K227 of LmjCS being involved in the complex forma... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2283096 Wed, 18 Mar 2009 04:00:00 +0100 2283096 http://www.medworm.com/index.php?rid=2236724&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20082421 Lipid hydroperoxides (LOOHs) in oxidized LDL (oxLDL) are potentially atherogenic compounds. Recently, H2S was identified as the third endogenous gasotransmitter in the vasculature. Hydrogen peroxide is known to be destroyed by H2S. Assuming that H2S may also react with LOOHs, the results show that H2S can destroy LOOHs in oxLDL. LOOH-enriched LDL after H2S-pretreatment was not further oxidized by endothelial cells and macrophages and the ability of oxLDL to induce HO-1 in endothelial cells was abolished by H2S pretreatment. HPLC analysis showed that 9-HPODE (9-hydroperoxy-octadecadienoic acid), a compound found in oxLDL, was reduced to 9-HODE (9-hydroxy-octadecadienoic acid) in presence of H2S. Thus, H2S may act as an antiatherogenic agent by reducing LOOHs to the less reactive LOHs and co... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2236724 Thu, 05 Mar 2009 05:00:00 +0100 2236724 http://www.medworm.com/index.php?rid=2213316&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081899 The c-Jun N-terminal kinase 1 (JNK1) plays a crucial role in the regulation of obesity-induced insulin resistance and is implicated in the pathology of type II diabetes. Its partner, JNK interacting protein 1 (JIP1), serves a scaffolding function facilitating JNK1 activation by mitogen-activated protein (MAP) kinase kinase 4 (MKK4) and MAP kinase kinase 7 (MKK7). For example, reduced insulin resistance and JNK activation are observed in JIP1 deficient mice. Based on the in vivo efficacy of a cell-permeable JIP peptide, the JIP/JNK interaction appears to be a potential target for JNK inhibition. The goal of this study was to identify small molecule inhibitors that disrupt the JIP/JNK interaction as an alternative approach to JNK inhibition. High throughput screening (HTS) was performed util... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2213316 Wed, 25 Feb 2009 05:00:00 +0100 2213316 http://www.medworm.com/index.php?rid=2203123&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20090082 Invasion-promoting membrane type-1 matrix metalloproteinase (MT1-MMP) is a key element in cell migration processes. To identify the proteins, which interact and, therefore, co-precipitate with this proteinase from cancer cells, we used the proteolytically potent wild-type (WT), the catalytically inert E240A (E240A) and the C-end truncated (tailless; ΔCT) MT1-MMP-FLAG constructs as baits. The identity of the pulled-down proteins was determined by LC-MS/MS and then confirmed with specific antibodies. We determined that in breast carcinoma MCF-7 cells ADP/ATP nucleoside transporter (ANT) efficiently interacted with the WT, E240A and ΔCT constructs. The WT and E240A constructs interacted also with alpha-tubulin, an essential component of the clathrin-mediated endocytosis. In turn... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2203123 Fri, 20 Feb 2009 05:00:00 +0100 2203123 http://www.medworm.com/index.php?rid=2196919&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20082222 In conclusion, 1 mM of butyrate and 1-15 mM of propionate increase MUC2 expression. The effects of butyrate on MUC2 mRNA are mediated via AP-1 and acetylation/methylation of histones at the MUC2 promoter. (Source: BJ Disease) BJ Disease journals http://www.medworm.com/rss/comments.php?id=2196919 Thu, 19 Feb 2009 05:00:00 +0100 2196919 http://www.medworm.com/index.php?rid=2179402&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081888 We report here the development of a mouse wherein the LPP1 gene (Ppap2a) was disrupted. The homozygous mice, which are phenotypically unremarkable, generally lack LPP1 mRNA and multiple tissues exhibit a substantial (35 – 95%) reduction in LPA phosphatase activity. Compared to wild type littermates, Ppap2atr/tr animals have increased levels of plasma LPA and LPA injected intravenously is metabolized at a four-fold slower rate. Our results demonstrate that LPA is rapidly metabolized in the bloodstream and that LPP1 is an important determinant of this turnover. These results indicate that LPP1 is a catabolic enzyme for LPA in vivo. (Source: BJ Disease) BJ Disease journals http://www.medworm.com/rss/comments.php?id=2179402 Thu, 12 Feb 2009 05:00:00 +0100 2179402 http://www.medworm.com/index.php?rid=2162058&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081787 Human African trypanosomiasis (HAT), caused by the protozoan parasite Trypanosoma brucei, is an emerging disease for which new drugs are needed. Expression of plasma membrane proteins, (e.g., variant surface glycoprotein (VSG)), is crucial for establishment and maintenance of an infection by T. brucei. Transport of a majority of proteins to the plasma membrane involves their translocation into the endoplasmic reticulum (ER). Thus, inhibition of protein import into the ER of T. brucei would be a logical target for discovery of lead compounds against trypanosomes. We have developed a trypanosome microsomal system (TbRM) that imports VSG_117 post-translationally. Using this system, MAL3-101, equisetin and CJ-21,058 were discovered as small molecule inhibitors of VSG_117 translocation into the... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2162058 Thu, 05 Feb 2009 05:00:00 +0100 2162058 http://www.medworm.com/index.php?rid=2158544&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20082240 Hemochromatosis is a genetic disorder of iron overload resulting from loss-of-function mutations in genes coding for the iron-regulatory proteins HFE, transferrin receptor 2, ferroportin, hepcidin, and hemojuvelin (HJV). Expression of the first four genes in retina has been established. Here we report on the expression of HJV. Since infection of retina with cytomegalovirus (CMV) causes blindness, we also investigated the expression of HJV and other iron-regulatory proteins in retina during CMV infection. Hjv mRNA was expressed in RPE/eyecup and neural retina in mouse. In situ hybridization and immunohistochemistry confirmed the presence of Hjv mRNA and protein in retinal pigment epithelium (RPE), outer and inner nuclear layers, and ganglion cell layer. Immunocytochemistry with cell lines a... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2158544 Wed, 04 Feb 2009 05:00:00 +0100 2158544 http://www.medworm.com/index.php?rid=2139504&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081351 Staphylococcus aureus is a dangerous human pathogen which antibiotic resistance is steadily increasing and no efficient vaccine is as yet available. This serious threat drives extensive studies on staphylococcal physiology and pathogenicity pathways, especially virulence factors. Serine protease like proteins (Spl) encoded by an operon containing up to six genes are a good example of poorly characterized secreted proteins likely involved in virulence. Here we describe an efficient heterologous expression system for SplA and detailed biochemical and structural characterization of the recombinant SplA protease. The enzyme shares a significant sequence homology to V8 protease and epidermolytic toxins which are well documented staphylococcal virulence factors. SplA has a very narrow substrate ... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2139504 Wed, 28 Jan 2009 05:00:00 +0100 2139504 http://www.medworm.com/index.php?rid=2136001&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081359 Wilson disease ATPase (ATP7B) has been implicated in the resistance of cancer cells to cisplatin. Using a simple in vivo assay in bacterial culture, we demonstrate that ATP7B can confer resistance to cisplatin by sequestering the drug in its N-terminal metal-binding domain without active drug extrusion from the cell. Expression of a protein fragment containing four N-terminal metal binding repeats of ATP7B (MBR1-4) protects cells from the toxic effects of cisplatin. One MBR1-4 molecule binds up to three cisplatin molecules at the copper-binding sites in the metal-binding repeats. These findings suggest that suppressing enzymatic activity of ATP7B may not be an effective way of combating cisplatin resistance. Rather, the efforts should be directed at preventing cisplatin binding to the prot... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2136001 Tue, 27 Jan 2009 05:00:00 +0100 2136001 http://www.medworm.com/index.php?rid=2136000&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20082368 Hepatic glucokinase (GK) plays a key role in maintaining glucose homeostasis. Many stimuli regulate GK activity by controlling its gene transcription. We hypothesized that endogenous lipophilic molecules modulate hepatic Gck expression. Lipophilic molecules were extracted from rat livers, saponified and re-constituted as a lipophilic extract (LE). LE synergized with insulin to induce primary hepatocyte, but not beta-cell, Gck expression in a sterol regulatory element-binding protein 1c (SREBP-1c) independent manner. The dramatic induction of Gck mRNA resulted in a significant increase of GK activity. Subsequently, the active molecules were identified as retinol and retinal by mass spectrometry after purification of the active LE fractions. Retinoids synergized with insulin to induce Gck ex... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2136000 Tue, 27 Jan 2009 05:00:00 +0100 2136000 http://www.medworm.com/index.php?rid=2109092&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20082068 Active site inhibitors of human immunodeficiency type 1 (HIV-1) protease (PR) block viral replication by preventing viral maturation. However, HIV-1 often develops resistance to active site inhibitors through multiple mutations in PR and therefore recent efforts have focused on inhibiting PR dimerization as an alternative approach. Dimerization inhibitors have been identified using kinetic analysis but additional characterization of the effect of these inhibitors on PR by physical methods has been difficult. Here, we identified a multi-drug resistant HIV-1 PR (PRMDR) that was highly resistant to autoproteolysis. Using this PR and a novel size-exclusion chromatographic approach that incorporated fluorescence and mass spectrometry detection, we were able to demonstrate inhibition of dimeriza... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2109092 Fri, 16 Jan 2009 05:00:00 +0100 2109092 http://www.medworm.com/index.php?rid=2103010&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20082197 Fragile X related proteins form a family implicated in RNA metabolism. Their sequence is composed of conserved N-terminal and central regions which contain Tudor and KH domains and of a divergent C-terminus with motifs rich in Arg and Gly. The most interesting member of the family is probably FMRP, since absence or mutation of this protein in human causes fragile X syndrome, the most common cause of inherited mental retardation. Understanding the structural properties of FMRP is essential for correlating it to its functions. The structures of isolated domains of FMRP have been reported, but nothing is yet known about the spatial arrangement of the different modules, partly because of difficulties in producing both the full-length protein and its multi-domain fragments in quantities, puriti... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2103010 Wed, 14 Jan 2009 05:00:00 +0100 2103010 http://www.medworm.com/index.php?rid=2082331&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081639 Chondroitin sulfate (CS) has been implicated in a variety of biological processes during development. Its biological functions are closely associated with characteristic sulfated structures. Here, we report the characterization of a zebrafish counterpart of chondroitin 4-O-sulfotransferase-1 (C4ST-1) and its functional importance in embryogenesis. Recombinant C4ST-1 showed a substrate preference for chondroitin and catalyzed the 4-O-sulfation of GalNAc residues, a highly frequent modification of CS in the embryos of zebrafish as well as other vertebrates. Whole-mount in situ hybridization revealed that C4ST-1 showed a distinct spatiotemporal expression pattern in the developing zebrafish embryo. During the segmentation stages, strong expression was observed along the body axis including th... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2082331 Tue, 06 Jan 2009 05:00:00 +0100 2082331 http://www.medworm.com/index.php?rid=2057516&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081526 Peroxiredoxin (Prx) is a multifunctional redox protein with thioredoxin-dependent peroxidase activity. Prx4 is present as a secretory protein in most tissues, whereas in sexually mature testes it is anchored in the endoplasmic reticulum (ER) membrane of spermatogenic cells via an uncleaved N-terminal hydrophobic peptide. We generated a Prx4-knockout mouse to investigate the function of Prx4 in vivo. Prx4−/y mice lacking Prx4 expression in all cells were obtained by mating Prx4flox/+ female mice with Cre-transgenic male mice that ubiquitously expressed Cre recombinase. The resulting Prx4−/y male mice were fertile, and most organs were nearly normal in size, except for testicular atrophy. The number of deoxynucleotidyl transferase-mediated dUTP nick end labeling-positive sperma... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2057516 Tue, 23 Dec 2008 05:00:00 +0100 2057516 http://www.medworm.com/index.php?rid=2057515&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081505 Antimicrobial polypeptides including lysozymes (Ly) have membrane perturbing activity and are well documented effector molecules of innate immunity. In cystic fibrosis, a hereditary disease with frequent lung infection with Pseudomonas aeruginosa, the free fatty acid docosahexaenoic acid (DA), but not oleic acid (OA), is decreased and DA supplementation has been shown to improve the clinical condition in these patients. We hypothesized that DA may alone, or in conjunction with Ly, exert antibacterial action against P. aeruginosa. We found that DA and Ly synergistically inhibit the metabolic activity of P. aeruginosa, in contrast to OA. Electron microscopy and equilibrium dialysis suggest that DA accumulates in the bacterial membrane in the presence of Ly. Surface plasmon resonance with liv... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2057515 Tue, 23 Dec 2008 05:00:00 +0100 2057515 http://www.medworm.com/index.php?rid=2057517&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081469 In this study, we examined the effects of gingipains on osteoclast differentiation. In co-cultures of mouse bone marrow cells and osteoblasts, formation of multinucleated osteoclasts induced by 1α,25-dihydroxyvitamin D3 [1α,25(OH)2D3] was augmented by Kgp but not by RgpB. A physiological concentration (0.1 nM) of 1α,25(OH)2D3 induced the osteoclast formation in the presence of 100 nM Kgp to the extent comparable to that induced by 10 nM 1α,25(OH)2D3. Kgp also enhanced osteoclastogenesis induced by various microbial components including lipopolysaccharide. Combined use of Kgp and 1α,25(OH)2D3 or lipopolysaccharide also increased the number of resorption pits developed on dentin slices, indicating the osteoclasts formed in the presence of Kgp possess bone-r... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2057517 Mon, 22 Dec 2008 05:00:00 +0100 2057517 http://www.medworm.com/index.php?rid=2037078&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20082097 In Ewing’s sarcomas, chromosomal translocations cause the N-terminal domain of the Ewing’s sarcoma protein (EWS) to fuse with the DNA-binding domains of the Ets family of transcription factors. Here we show that EWS and EWS-Fli1, the fusion most frequently found in Ewing’s sarcomas, become phosphorylated at Thr79 in response to either mitogens or DNA damaging agents. The much weaker mitogen-induced phosphorylation of EWS is catalysed by the MAP kinases ERK1 and ERK2, while the much stronger phosphorylation of EWS induced by the DNA alkylating agent methyl methanesulphonate (MMS) can be catalysed by JNK and at least one other protein kinase distinct from ERK1/ERK2. In contrast, the phosphorylation of EWS-Fli1 induced by MMS was largely mediated by p38α/p38&#x03B2... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2037078 Tue, 16 Dec 2008 05:00:00 +0100 2037078 http://www.medworm.com/index.php?rid=2037077&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081983 In the blood plasma of humans and rats, ceruloplasmin is the major Cu-binding protein and ferroxidase, accounting for 70% of the copper, the rest binding primarily to albumin and a macroglobulin. Systematic studies with fresh plasma were carried out to compare what occurs in the mouse. C57BL6 mice had half as much copper and p-phenylene diamine (pPD) oxidase activity as humans and rats; 20-40% as much ferroxidase activity as humans (by three different assays); and less inhibition by azide. Plasma of ceruloplasmin knockout mice had no pPD oxidase activity, but retained >50% ferroxidase activity (not as affected by azide). Modeling of mouse ceruloplasmin against the known x-ray structure of the human indicated subtle but potentially significant changes in the pPD and azide binding sites. Pur... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2037077 Tue, 16 Dec 2008 05:00:00 +0100 2037077 http://www.medworm.com/index.php?rid=2037080&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081874 Antizymes are small polyamine-induced proteins that function as feedback regulators of cellular polyamine homeostasis. They bind to transient ornithine decarboxylase (ODC) monomeric subunits, resulting in inhibition of ODC activity and targeting ODC to ubiquitin-independent proteasomal degradation. Antizyme 3 (Az3) is a mammalian antizyme isoform expressed exclusively in testicular germ cells and therefore considered as a potential regulator of polyamines during spermatogenesis. We show here that unlike antizyme 1 (Az1) and antizyme 2 (Az2), which efficiently inhibit ODC activity and stimulate its proteasomal degradation, antizyme 3 poorly inhibits ODC activity and fails to promote ODC degradation. Furthermore, Az3 actually stabilizes ODC, probably by protecting it from the effect of Az1. ... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2037080 Mon, 15 Dec 2008 05:00:00 +0100 2037080 http://www.medworm.com/index.php?rid=2037079&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081448 Postprandial blood glucose clearance is mediated by glucose transporter isoform 4 (Glut4) which is translocated from an intracellular storage pool to the plasma membrane in response to insulin. The nature of the intracellular storage pool of Glut4 is not well understood. Immunofluorescence staining shows that, under basal conditions, the major population of Glut4 resides in the perinuclear compartment. At the same time, biochemical fractionation reveals that Glut4 is localized in insulin-responsive vesicles, or IRVs. The relationship between the perinuclear Glut4 compartment and the IRVs is not known. Here, we have exchanged the C-termini of Glut4 and cellugyrin, another vesicular protein that is not localized in the IRVs and has no insulin response. We have found that Glut4 with the cellu... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2037079 Mon, 15 Dec 2008 05:00:00 +0100 2037079 http://www.medworm.com/index.php?rid=2009024&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081869 Recent studies suggest that immature, core-glycosylated ΔF508-CFTR (the predominant mutant form of the Cystic Fibrosis Transmembrane conductance Regulator) can reach the plasma membrane under some conditions. We investigated this possibility since it has implications for understanding how therapeutics rescue the trafficking of mutant CFTR and perhaps other misfolded proteins. Core-glycosylated CFTR was labelled and pulled down on streptavidin beads after exposure to sulfo-NHS-SS-biotin (Mr = 606.7) however intracellular proteins were also detected in the precipitates. When the R domain of CFTR was expressed in the cytosol of BHK cells as a soluble polypeptide it was also labelled after surface biotinylation and pulled down on streptavidin beads. Intracellular biotinylation was reduc... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2009024 Thu, 04 Dec 2008 05:00:00 +0100 2009024 http://www.medworm.com/index.php?rid=2009023&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20082176 This study compares the major insulin signalling pathways in skeletal muscle isolated from PCOS and controls. We measured whole body insulin sensitivity and insulin signalling in skeletal muscle biopsies taken before and after acute exposure to hyperinsulinaemia in nine women diagnosed with PCOS and seven controls. We examined the expression, basal activity and response to in vivo insulin stimulation of three signalling molecules within these human muscle samples, namely, insulin receptor substrate-1 (IRS1), protein kinase-B (PKB) and extracellular regulated protein kinase (ERK1/ERK2). There was no significant difference in the expression, basal activity, or activation of IRS1 or PKB between PCOS and control subjects. However, there was a severe attenuation of insulin stimulation of the ER... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2009023 Thu, 04 Dec 2008 05:00:00 +0100 2009023 http://www.medworm.com/index.php?rid=2009025&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081793 The expression of the β-amyloid precursor protein (APP), which plays a key role in the development of Alzheimer's disease, is regulated by a variety of cellular mediators in a cell-dependent manner. In the present study, we present evidence that p53 regulates the expression of the APP gene in neuroblastoma cells. Transient expression of ectopic p53, as well as the activation of endogenous p53 by the DNA-damaging drug camptothecin or MDM2 depletion, decrease intracellular levels of APP in N2aβ neuroblastoma cells. This effect was also observed in primary cultures of rat neurons as well as in SH-SY5Y cells, a human neuroblastoma cell line. Transient transfection studies using plasmids which contain progressive deletions of the 5' region of the gene, demonstrate that p53 repress... BJ Disease journals http://www.medworm.com/rss/comments.php?id=2009025 Wed, 03 Dec 2008 05:00:00 +0100 2009025 http://www.medworm.com/index.php?rid=1986823&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081765 We have developed a polypeptide FRET substrate (MV11F) for the endopeptidase activity of lysostaphin. Site-directed mutants of lysostaphin that abolished the killing activity against Staphylococcus aureus also completely inhibited the endopeptidase activity against the MV11 FRET substrate. Lysostaphin-producing staphylococci are resistant to killing by lysostaphin through incorporation of serine residues at position 3 and 5 of the pentaglycine cross-bridge in their cell walls. The MV11 FRET substrate was engineered to introduce a serine residue in turn at four positions of the pentaglycine target site and revealed that only a serine residue at position 3 completely inhibited cleavage. The introduction of random, natural amino acid substitutions at position 3 of the pentaglycine target site... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1986823 Tue, 25 Nov 2008 05:00:00 +0100 1986823 http://www.medworm.com/index.php?rid=1965945&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20082011 In Mycobacterium tuberculosis the genes hsaD (2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoic acid hydrolase) and nat (arylamine N-acetyltransferase) are essential for survival inside of host macrophages. These genes act as an operon and have been suggested to be involved in cholesterol metabolism. However, the role of NAT in this catabolic pathway has not been determined. In an effort to better understand the function of these proteins, we have expressed, purified and characterized TBNAT and HsaD from M. tuberculosis. Both proteins demonstrated remarkable heat stability with TBNAT and HsaD retaining >95% of their activity following incubation at 60ºC for 30 min. The first and second domain of TBNAT was demonstrated to be very important to the heat stability of the protein, as the transfer... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1965945 Tue, 18 Nov 2008 05:00:00 +0100 1965945 http://www.medworm.com/index.php?rid=1957331&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081572 α-Synuclein is a pathological component of Parkinson's disease (PD) by participating in the Lewy body formation. JC-1 has been shown to interact with α-synuclein at the acidic C-terminal region with Kd of 2.6 μM. JC-1 has discriminated the fibrillation states of α-synuclein - monomeric, oligomeric intermediate, and fibrillar forms - by emitting the enhanced binding fluorescence of different colors at 590 nm, 560 nm, and 538 nm, respectively, with the common excitation at 490 nm. The fibrillation state-specific interaction of JC-1 allowed us to perform real-time analyses of the α-synuclein fibrillation in the presence of iron as a fibrillation inducer, rifampicin as a fibrillation inhibitor, baicalein as a defibrillation agent, and dequalinium as a protofi... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1957331 Fri, 14 Nov 2008 05:00:00 +0100 1957331 http://www.medworm.com/index.php?rid=1949200&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081541 Prions are infectious proteins responsible for a group of fatal neurodegenerative diseases called transmissible spongiform encephalopathies (TSEs), or prion diseases. In mammals, prions reproduce themselves by recruiting the normal cellular protein (PrPC) and inducing its conversion to the disease-causing isoform denominated PrPSc. Recently, anti-prion antibodies have been shown to permanently cure prion-infected cells. However, the inability of full-lenght antibodies and proteins to cross the blood-brain barrier (BBB) hampers their use in the therapy of TSEs in vivo. Alternatively, brain delivery of prion-specific scFv by adeno-associated virus (AAV) transfer delays the onset of the disease in infected mice, although protection is not complete. We investigated the anti-prion effects of a... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1949200 Tue, 11 Nov 2008 05:00:00 +0100 1949200 http://www.medworm.com/index.php?rid=1935977&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081848 Human pancreatic trypsinogens undergo post-translational sulfation on Tyr154, catalyzed by the Golgi-resident enzyme tyrosylprotein sulfotransferase 2. Sequence alignments suggest that sulfation of Tyr154 is facilitated by a unique sequence context characteristically found in primate trypsinogens. In search for genetic variants that might alter this sulfation motif, we identified a single nucleotide polymorphism (c.457G>C) in the human anionic trypsinogen gene (PRSS2), which changed Asp153 to His (p.D153H). The p.D153H variant is common in subjects of African origin, with a minor allele frequency of 9.2%, whereas it is absent in subjects of European descent. We demonstrate that Asp153 is the main determinant of tyrosine sulfation in anionic trypsinogen, as both the natural p.D153H variatio... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1935977 Thu, 06 Nov 2008 05:00:00 +0100 1935977 http://www.medworm.com/index.php?rid=1898019&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081424 Cystic Fibrosis is caused by mutations in CFTR which cause its mistrafficking and/or dysfunction as a regulated chloride channel on the apical surface of epithelia. CFTR is a member of the ATP Binding Cassette (ABC) superfamily of membrane proteins and a disease-causing missense mutation within the ABC signature sequence; G551D-CFTR, exhibits defective phosphorylation and ATP dependent channel gating. Studies of the purified and reconstituted G551D-CFTR protein revealed that faulty gating is associated with defective ATP binding and ATPase activity, reflecting the key role for G551 in these functions. Recently, high-throughput screens of chemical libraries led to identification of modulators which enhance channel activity of G551D-CFTR. However, the molecular target (s) for these modulator... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1898019 Thu, 23 Oct 2008 04:00:00 +0100 1898019 http://www.medworm.com/index.php?rid=1884871&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081241 CD22 (Siglec-2), a negative regulator of B cell signaling, binds to α2,6 sialic acid-linked glycoconjugates including a sialyl-Tn antigen that is one of the typical tumor-associated carbohydrate antigens expressed on various mucins. Many epithelial tumors secrete mucins into tissues and/or the bloodstream. Mouse mammary adenocarcinoma cells, TA3-Ha, produce a mucin named epiglycanin but a subline of them, TA3-St, does not. Epiglycanin binds to CD22 and inhibits B cell signaling in vitro. The in vivo effect of mucins in the tumor-bearing state was investigated using these cell lines. It should be noted that splenic marginal zone B cells were dramatically reduced in the mice bearing TA3-Ha cells but not in those bearing TA3-St cells, this being consistent with the finding that the thy... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1884871 Fri, 17 Oct 2008 04:00:00 +0100 1884871 http://www.medworm.com/index.php?rid=1835625&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081134 This study illuminates new properties of curcumin and increases the evidence that this dietary polyphenol can be exploited for rational drug design in human leishmaniasis. (Source: BJ Disease) BJ Disease journals http://www.medworm.com/rss/comments.php?id=1835625 Mon, 29 Sep 2008 04:00:00 +0100 1835625 http://www.medworm.com/index.php?rid=1831428&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080744 Elevated plasma concentrations of lipoprotein(a) [Lp(a)] are an emerging risk factor for atherothrombotic disease. Apolipoprotein(a) [apo(a)], the unique glycoprotein component of Lp(a), contains tandem repeats of a plasminogen kringle (K) IV-like domain. In light of recent studies suggesting that apo(a)/Lp(a) affects endothelial function, we evaluated the effects of apo(a)/Lp(a) on growth and migration of cultured human umbilical vein endothelial cells (HUVECs). Two full-length recombinant apo(a) [r-apo(a)] variants (12K and 17K), as well as Lp(a), were able to stimulate HUVEC growth and migration to a comparable extent; 17K r-apo(a) also decreased the levels of total and active transforming growth factor-β secreted by these cells. Using additional r-apo(a) variants corresponding t... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1831428 Fri, 26 Sep 2008 04:00:00 +0100 1831428 http://www.medworm.com/index.php?rid=1804413&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081615 In this study, we examined HNE metabolism in isolated aerobic and ischemic rat hearts. In aerobic hearts, reagent [3H]-HNE was glutathiolated, oxidized to [3H]-4-hydroxynonenoic acid, and reduced to [3H]-1,4-dihydroxynonene. In ischemic hearts, [3H]-4-hydroxynonenoic acid formation was inhibited and higher levels of [3H]-1,4-dihydroxynonene and [3H]-glutathionyl-HNE were generated. Metabolism of [3H]-HNE to [3H]-4-hydroxynonenoic acid was restored upon reperfusion. Reperfused hearts were more efficient at metabolizing HNE than non-ischemic hearts. Ischemia increased the myocardial levels of endogenous HNE and 1,4-dihydroxynonene but not 4-hydroxynonenoic acid. Isolated cardiac mitochondria metabolized [3H]-HNE primarily to [3H]-4-hydroxynonenoic acid and minimally to [3H]-1,4-dihydroxynon... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1804413 Thu, 18 Sep 2008 04:00:00 +0100 1804413 http://www.medworm.com/index.php?rid=1787242&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081132 Human colon cancer cells and primary colon cancer silence the gene coding for lactate dehydrogenase LDH-B and upregulate the gene coding for lactate dehydrogenase LDH-A, resulting in effective conversion of pyruvate into lactate. This is associated with markedly reduced levels of pyruvate in cancer cells compared to non-malignant cells. The silencing of LDH-B in cancer cells occurs via DNA methylation, with involvement of DNA methyltransferases DNMT1 and DNMT3b. Colon cancer is also associated with the expression of pyruvate kinase M2, a splice variant with low catalytic activity. We have shown recently that pyruvate is an inhibitor of histone deacetylases (HDACs). Here we show that pyruvate is a specific inhibitor of HDAC1 and HDAC3. Lactate has no effect on any of the HDACs examined. Col... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1787242 Fri, 12 Sep 2008 04:00:00 +0100 1787242 http://www.medworm.com/index.php?rid=1758240&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081421 We have recently demonstrated that 5-chloro-7-iodo-8-hydroxyquinoline (clioquinol) acts as a zinc ionophore and induces apoptosis of human cancer cells. However, the mechanisms of clioquinol/zinc-induced apoptotic cell death remain to be further elucidated. Using fluorescence-labeled probes, the present study has examined intracellular zinc distribution after clioquinol treatment in human cancer cells in order to identify cellular targets for zinc ionophores. DU 145, a human prostate cancer line, was chosen as a model system for this study, and results were confirmed in other human cancer cell lines. Although treatment of cancer cells with 50 µM zinc chloride for three days had no effect on cell viability, addition of clioquinol dramatically enhanced the cytotoxicity, confirming our... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1758240 Wed, 03 Sep 2008 04:00:00 +0100 1758240 http://www.medworm.com/index.php?rid=1736583&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080868 Nonalcoholic fatty liver disease (NAFLD) associated with obesity and the cardiometabolic syndrome is an important medical problem affecting up to 20% of western populations. Evidence indicates that mitochondrial dysfunction plays a critical role in NAFLD initiation and progression to the more serious condition of nonalcoholic steatohepatitis (NASH). Herein, we hypothesize that mitochondrial defects induced by exposure to a high fat diet (HFD) contribute to a hypoxic state in liver and this is associated with increased protein modification by reactive nitrogen species (RNS). To test this concept, C57BL/6 mice were pair-fed a control diet and HFD containing 35% and 71% total calories from fat, respectively, for 8 or 16 weeks and liver hypoxia, mitochondrial bioenergetics, nitric oxide (NO)-d... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1736583 Thu, 28 Aug 2008 04:00:00 +0100 1736583 http://www.medworm.com/index.php?rid=1731703&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081276 The role of the functional architecture of the human acetylcholinesterase (HuAChE) in reactivity toward the carbamates pyridostigmine, rivastigmine and several analogs of physostigmine, that are currently used or considered for use as drugs for Alzheimer’s disease, was analysed using over 20 mutants of residues that constitute the interaction subsites in the active center. Both steps of the HuAChE carbamylation reaction, formation of the Michaelis complex as well as the nucleophilic process, are sensitive to accommodation of the ligand by the enzyme. For certain carbamate - HuAChE combinations, the mode of inhibition shifted from a covalent to a noncovalent type, according to the balance between dissociation and covalent reaction rates. Whereas the charged moieties of pyridostigmin... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1731703 Tue, 26 Aug 2008 04:00:00 +0100 1731703 http://www.medworm.com/index.php?rid=1715702&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080918 In vitro and in vivo studies have demonstrated that unconjugated bilirubin (UCB) is neurotoxic. Although previous studies suggested that both MRP1 and MDR1 proteins can protect cells against accumulation of UCB, direct comparison of their role in UCB transport was never performed. To this end, we modulated expression of MRP1 and MDR1 in human neuroblastoma SH-SY5Y cells using an inducible siRNA expression system. The effects of in vitro exposure to clinically-relevant levels of UCB were compared between cells with similar reductions in the expression of MRP1 or MDR1; cells expressing endogenous levels of the two proteins were used as controls. MRP1 deficient cells accumulated a significantly higher amount of UCB, while UCB uptake of MDR1 deficient cells was comparable to controls. Cell fun... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1715702 Tue, 19 Aug 2008 04:00:00 +0100 1715702 http://www.medworm.com/index.php?rid=1672832&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081224 The effect of COX-2-dependent prostaglandins (PGs) in acute liver injury has been investigated in transgenic mice that express human COX-2 in hepatocytes. We have used three well established models of liver injury: Lipopolysaccharide injury in D-galactosamine preconditioned mice (LPS/D-GalN), the hepatitis induced by concanavalin A (ConA) and the proliferation of hepatocytes in regenerating liver after partial hepatectomy (PH). Our data demonstrate that PGs synthesis decreases in hepatocytes the susceptibility to LPS/D-GalN or ConA-induced liver injury as deduced by significant lower levels of the pro-inflammatory profile and plasmatic aminotransferases in transgenic mice, an effect suppressed by COX-2 selective inhibitors. These transgenic animals express higher levels of anti-apoptot... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1672832 Fri, 01 Aug 2008 04:00:00 +0100 1672832 http://www.medworm.com/index.php?rid=1644165&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080054 Discussion: These results show that RAGE and PI3-Kinase modulate adhesion and migration rate of neutrophils on AGE-Collagen. Modulation of adhesiveness may account for the change in neutrophil migration rate on AGE-collagen. Since neutrophils rely on their ability to move to perform their function as the first line of defense against bacterial invasion, glycoxidation of collagen may participate in the suppression of normal host defense in diabetic and uremic patients. (Source: BJ Disease) BJ Disease journals http://www.medworm.com/rss/comments.php?id=1644165 Tue, 22 Jul 2008 04:00:00 +0100 1644165 http://www.medworm.com/index.php?rid=1644164&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080981 The newly synthesized surfactant protein C precursor (proSP-C) is an integral endoplasmic reticulum (ER) membrane protein with a single metastable polyVal α-helical transmembrane domain that comprises two thirds of the mature peptide. More than 20 mutations in the ER-lumenal, C-terminal domain of proSP-C (CTC), are associated with interstitial lung disease (ILD), and some of the mutations cause intracellular accumulation of cytotoxic protein aggregates and a corresponding decrease in mature SP-C. Here it is shown that (i) human embryonic kidney cells expressing the ILD associated mutants proSP-CL188Q and proSP-CΔExon4 accumulate Congo red positive amyloid-like inclusions, while cells transfected with the mutant proSP-CI73T do not, (ii) transfection of CTC into cells expressin... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1644164 Tue, 22 Jul 2008 04:00:00 +0100 1644164 http://www.medworm.com/index.php?rid=1594745&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20081068 ATP binding to the first (NBD1) and second (NBD2) nucleotide binding sites of CFTR are divalent cation independent and dependent steps, respectively (Aleksandrov et al, J Biol Chem 277, 15419-25, 2002). Subsequent to the initial binding, Mg2+ drives rapid hydrolysis at the second site while promoting non-exchangeable trapping of the nucleotide at the first site. This occlusion at the first site of functional wild-type CFTR is somewhat similar to that which occurs when the catalytic glutamates in both of the hydrolytic sites of P-glycoprotein are mutated, which has been proposed to be the result of dimerization of the two NBDs and represent a transient intermediate formed during ATP hydrolysis (Tombline and Senior, J Bioenerg Biomembr 37, 497-500, 2005). To test the possible relevance of th... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594745 Tue, 08 Jul 2008 04:00:00 +0100 1594745 http://www.medworm.com/index.php?rid=1594746&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080398 A novel gene family coding for putative antimicrobial peptides was identified in the EST data base of the sea squirt Ciona intestinalis, and one of these genes was molecularly cloned from the Northern European Ciona subspecies. In situ hybridisation and immunocytochemical analysis revealed that the natural peptide is synthesized and stored in a distinct hemocyte type, the univacuolar non-refractile granulocytes. By semiquantitative RT-PCR analysis it was shown that the expression of the gene is markedly upregulated in hemocytes after immune challenge. To evaluate the antimicrobial potency of the putative defense protein, we synthesized a peptide corresponding to its cationic core region. The peptide was highly effective against Gram-negative and Gram-positive bacteria including several hum... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594746 Fri, 04 Jul 2008 04:00:00 +0100 1594746 http://www.medworm.com/index.php?rid=1594747&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080666 Proteins of the SIR2 family are characterized by a conserved catalytic domain that exerts unique NAD+-dependent deacetylase activity on histone and various other cellular substrates. Previous reports from our team have identified a Leishmania infantum gene encoding a cytosolic protein termed LiSIR2RP1 that belongs to the SIR2 family. Targeted disruption of one LiSIR2RP1 gene allele led to decreased amastigote virulence, in vitro as well as in vivo. In the present study, attempts were made for the first time to explore and characterize the LiSIR2RP1 enzymatic functions. The LiSIR2RP1 exhibited robust NAD+-dependent deacetylase and ADP-ribosyltransferase activities. Moreover, LiSIR2RP1 is capable of deacetylating tubulin, either in dimers or, when present, in taxol-stabilized microtubules or... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594747 Thu, 03 Jul 2008 04:00:00 +0100 1594747 http://www.medworm.com/index.php?rid=1594748&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080965 In this study, we used site-directed mutagenesis to identify residues affecting the substrate specificity of human ERAP-1 and identified Gln-181 as important for enzymatic activity and substrate specificity. Replacement of Gln-181 with Asp resulted in a significant change in substrate specificity with Q181D ERAP-1 showing a preference for basic amino acids. In addition, Q181D ERAP-1 cleaved natural peptides possessing a basic amino acid of the N-terminal end more efficiently than did the wild-type enzyme, whereas its cleavage of peptides with a non-basic amino acid was significantly reduced. Another mutant enzyme, Q181E, also revealed some preference for peptides with a basic N-terminal amino acid although it had little hydrolytic activity toward synthetic peptides tested. Other mutant enz... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594748 Tue, 01 Jul 2008 04:00:00 +0100 1594748 http://www.medworm.com/index.php?rid=1594749&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080328 Two of the most common signaling pathways in breast cancer are the ER ligand activation pathway and the E-cadherin-snai1-slug EMT pathway. Although these pathways have been thought to interact indirectly, the present study is the first to observe direct interactions between these pathways that involves the regulation of slug expression. Specifically we report that ligand-activated ERα suppressed slug expression directly by repression of transcription and that knockdown of ERα with RNA interference increased slug expression. More specifically, slug expression was downregulated in ERα negative MDA-MB-468 cells transfected with ERα after treatment with 17β-estradiol (E2). The downregulation of slug in the ER-α positive MCF-7 line was mediated by direc... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594749 Mon, 30 Jun 2008 04:00:00 +0100 1594749 http://www.medworm.com/index.php?rid=1594750&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080411 Late-infantile neuronal ceroid lipofuscinosis (LINCL) is a fatal neurodegenerative disease resulting from mutations in the gene encoding the lysosomal protease tripeptidyl-peptidase I (TPPI). TPPI is expressed ubiquitously throughout the body but disease appears restricted to the brain. One explanation for the absence of peripheral pathology is that in tissues other than brain, other proteases may compensate for the loss of TPPI. One such candidate is another lysosomal aminopeptidase, dipeptidyl-peptidase I (DPPI), which appears to have overlapping substrate specificity with TPPI and is expressed at relatively low levels in brain. Compensation for the loss of TPPI by DPPI may have therapeutic implications for LINCL and we have investigated this possibility using mouse genetic models. Our r... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594750 Fri, 20 Jun 2008 04:00:00 +0100 1594750 http://www.medworm.com/index.php?rid=1594751&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20071696 We report here an apparent relationship between altered cytokine expression and amyloid A accumulation in systemically inflamed tissues. The prevalence of SAA monomers and proteolytic oligomers in spleen AEF is consistent to suggest that extrahepatic SAA processing might lead to local accumulation of amyloidogenic proteins at the SAA production site. (Source: BJ Disease) BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594751 Wed, 18 Jun 2008 04:00:00 +0100 1594751 http://www.medworm.com/index.php?rid=1594752&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080234 Eukaryotic-type Serine/Threonine protein kinases in bacteria have been implicated in controlling a host of cellular activities. PknA is one of the eleven such protein kinases from Mycobacterium tuberculosis which regulates morphological changes associated with cell division. We provide here the evidence for the ability of PknA to transphosphorylate mycobacterial UDP-N-acetylmuramoyl-L-alanine:D-glutamate-ligase (mMurD), the enzyme involved in peptidoglycan biosynthesis. Its co-expression in Escherichia coli along with PknA resulted in phosphorylation of mMurD. Consistent with these observations, results of the solid phase binding assays revealed a high affinity in vitro binding between the two proteins. Furthermore, overexpression of m-murD in M. smegmatis yielded a phosphorylated protein.... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594752 Mon, 16 Jun 2008 04:00:00 +0100 1594752 http://www.medworm.com/index.php?rid=1594753&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080889 Trypanosoma brucei tryparedoxin-dependant peroxidase (TbTDPX) is a genetically validated drug target in the fight against African sleeping sickness. Despite its similarity to members of the glutathione peroxidase (GPX) family, TbTDPX2 is functional as a monomer, lacks a selenocysteine residue, relying instead on peroxidatic and resolving cysteines for catalysis and uses tryparedoxin rather than glutathione as electron donor. Kinetic studies indicate a saturable ping-pong mechanism, unlike selenium-dependent GPXs which display infinite Km and Vmax values. The structure of the reduced enzyme at 2.1 Å-resolution reveals that the catalytic thiol groups are widely separated (19 Å) and thus unable to form a disulphide bond without a large conformational change in the secondary stru... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594753 Wed, 04 Jun 2008 04:00:00 +0100 1594753 http://www.medworm.com/index.php?rid=1594755&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080489 Caffeine has long been used as a pharmacological probe for studying ryanodine receptor (RyR)-mediated Ca2+ release and cardiac arrhythmias. However, the precise mechanism by which caffeine activates RyRs is elusive. Here we investigated the effects of caffeine on spontaneous Ca2+ release and on the response of single cardiac RyR (RyR2) channels to luminal or cytosolic Ca2+. We found that HEK293 cells expressing RyR2 displayed partial or “quantal” Ca2+ release in response to repetitive additions of submaximal concentrations of caffeine. This quantal Ca2+ release was abolished by ryanodine. Monitoring of endoplasmic reticulum luminal Ca2+ revealed that caffeine reduced the luminal Ca2+ threshold at which spontaneous Ca2+ release occurs. Interestingly, spontaneous Ca2+ release i... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594755 Tue, 03 Jun 2008 04:00:00 +0100 1594755 http://www.medworm.com/index.php?rid=1594754&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080622 ATB0,+ (SLC6A14) is a Na+/Cl- -coupled amino acid transporter whose expression is upregulated in cancer. 1-Methyltryptophan is an inducer of immune surveillance against tumor cells through its ability to inhibit indoleamine dioxygenase. Here we investigated the role of ATB0,+ in the uptake of 1-methyltryptophan as a potential mechanism for entry of this putative anticancer drug into tumor cells. These studies show that 1-methyltryptophan is a transportable substrate for ATB0,+. The transport process is Na+/Cl- -dependent with a Na+:Cl-:1-methyltryptophan stoichiometry of 2:1:1. Evaluation of other derivatives of tryptophan has led to identification of α-methyltryptophan as a blocker, not a transportable substrate, for ATB0,+. ATB0,+ can transport 18 of the 20 proteinogenic amino aci... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594754 Tue, 03 Jun 2008 04:00:00 +0100 1594754 http://www.medworm.com/index.php?rid=1594756&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080639 The Phosphoinositide-3-kinase (PI3K) pathway regulates cell proliferation, survival and migration and is consequently of great interest for targeted cancer therapy. Using a panel of small molecule PI3K isoform-selective inhibitors in a diverse set of breast cancer cell lines, we demonstrate that the biochemical and biological responses were highly variable and dependent on the genetic alterations present. p110α inhibitors were generally effective in inhibiting the phosphorylation of Akt and S6, two downstream components of PI3K signaling, in most cell lines examined. In contrast, p110β selective inhibitors only reduced Akt phosphorylation in PTEN mutant cell lines, which was not accompanied by loss of S6 phosphorylation. PI3K inhibitors reduced cell viability by causing a cel... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594756 Thu, 22 May 2008 04:00:00 +0100 1594756 http://www.medworm.com/index.php?rid=1594757&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080435 Ethanolamine (EtN) and choline (Cho) are major components of the trypanosome membrane phospholipids, in the form of phosphatidylethanolamine (GPEtn) and phosphatidylcholine (GPCho). Ethanolamine is also found as an integral component of the glycosylphosphatidylinositol (GPI) anchor that is required for membrane attachment of cell surface proteins, most notably the variant surface glycoproteins. The de novo synthesis of GPEth and GPCho starts with the generation of phosphoethanolamine and phosphocholine by ethanolamine and choline kinases via the Kennedy pathway. Database mining revealed two putative choline/ethanolamine kinases (C/EKs) in the T. brucei genome, which were cloned, over-expressed, purified and characterised. TbEK1 was shown to be catalytically active as an ethanolamine-specif... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594757 Tue, 20 May 2008 04:00:00 +0100 1594757 http://www.medworm.com/index.php?rid=1594758&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080506 Cationic peptides, known to disrupt bacterial membranes, are being developed as promising agents for therapeutic intervention against infectious disease. Here, we investigate structure-activity relationships in the bacterial membrane disruptor βpep25, a peptide 33mer. For insight into which amino acid residues are functionally important, we synthesized alanine scanning variants of βpep25 and assessed their ability to kill bacteria (E. coli, P. aeruginosa, and S. aureus) and to neutralize lipopolysaccharide (LPS). Activity profiles were found to vary with the bacterial strain examined. Specific cationic and smaller hydrophobic alkyl residues were crucial to optimal bactericidal activity against the Gram negative bacteria, whereas larger hydrophobic and cationic residues mediat... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594758 Mon, 19 May 2008 04:00:00 +0100 1594758 http://www.medworm.com/index.php?rid=1594759&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080074 Human equilibrative nucleoside transporter 1 (hENT1) is inhibited by nanomolar concentrations of various structurally distinct coronary vasodilator drugs, including dipyridamole, dilazep, draflazine, soluflazine and nitrobenzylmercaptopurine ribonucleoside (NBMPR). When a library of randomly mutated hENT1 cDNAs was screened using a yeast-based functional complementation assay for resistance to dilazep, a clone containing the W29G mutation was identified. Multiple sequence alignments revealed that this residue was highly conserved. Mutations at W29 were generated and tested for adenosine transport activity and inhibitor sensitivity. W29 mutations significantly reduced the apparent Vmax and/or increased the apparent Km values for adenosine transport. W29 mutations increased the IC50 values f... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594759 Thu, 08 May 2008 04:00:00 +0100 1594759 http://www.medworm.com/index.php?rid=1594760&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080468 In this study, HOCl and HOBr are shown to react rapidly with macrophage (J774A.1) cells resulting in a greater extent of cell lysis compared to HOSCN. However, HOSCN induces apoptosis and necrosis with greater efficacy, and at lower concentrations than HOCl or HOBr. Apoptosis occurs in conjunction with an increased release of cytochrome c into the cytosol, but no associated increase in caspase activity. Similarly, apoptosis is observed on treating the cells in the presence of a caspase inhibitor, suggesting that it is mediated by a caspase-independent pathway. HOSCN oxidised protein thiols more efficiently than either HOCl or HOBr. The greater efficacy of HOSCN in inducing apoptosis, is attributed to selective damage to critical mitochondrial membrane protein thiol groups, resulting in inc... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594760 Wed, 07 May 2008 04:00:00 +0100 1594760 http://www.medworm.com/index.php?rid=1594761&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080406 Following its identification as a liver-expressed antimicrobial peptide, the hepcidin peptide was later shown to be a key player of in iron homeostasis. It is now proposed to be the “iron-hormone” which, by interacting with the iron transporter ferroportin, prevents further iron import into the circulatory system. This conclusion was reached using the corresponding synthetic peptide, emphasizing the functional importance of the mature 25-mer peptide, but omitting the possible functionality of its maturation. From urine-purified native hepcidin, we recently identified a proportion of iron-hepcidin complex. This interaction was further investigated by computer modeling and, based on sequence similarity with metallothionein, a 3D model of hepcidin containing one atom of iron was... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594761 Thu, 01 May 2008 04:00:00 +0100 1594761 http://www.medworm.com/index.php?rid=1594763&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080397 Human glutathione transferase A3-3 (hGST A3-3) efficiently catalyzes steroid Δ5-Δ4 double-bond isomerization in vitro, using glutathione as a cofactor. This chemical transformation is an obligatory reaction in the biosynthesis of steroid hormones and follows the oxidation of 3β-hydroxysteroids catalyzed by 3β-hydroxysteroid dehydrogenase (3β-HSD). The isomerization has commonly been ascribed to a supplementary function of 3β-HSD. The current study is the first to present evidence that hGST A3-3 contributes to this step in steroid hormone biosynthesis in complex cellular systems. First, we find glutathione-dependent Δ5-Δ4 isomerase activity in whole cell extracts prepared from human steroidogenic cells. Second, effective inhibitors of ... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594763 Tue, 22 Apr 2008 04:00:00 +0100 1594763 http://www.medworm.com/index.php?rid=1594762&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20080260 MUC5B is the predominant polymeric mucin in human saliva (Thornton, Khan, Mehrotra, Howard, Veerman, Packer, and Sheehan (1999) Glycobiology 9, 293-302) where it contributes to oral cavity hydration and protection. More recently the gene for another putative polymeric mucin, MUC19, has been shown to be expressed in human salivary glands (Chen, Zhao, Kalaslavadi, Hamati, Nehrke, Le, Ann, and Wu, (2004) Am.J.Respir.Cell Mol.Biol. 30, 155-165). However, to date, the MUC19 mucin has not been isolated from human saliva. Our aim was therefore to purify and characterize the MUC19 glycoprotein from human saliva. Saliva was solubilised in 4M guanidinium chloride and the high-density mucins were purified by density gradient centrifugation. The presence of MUC19 was investigated using tandem mass spe... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594762 Tue, 22 Apr 2008 04:00:00 +0100 1594762 http://www.medworm.com/index.php?rid=1594764&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20071690 Cystic fibrosis (CF) is most commonly caused by deletion of a residue (ΔF508) in the cystic fibrosis transmembrane conductance regulator (CFTR) protein. The misfolded mutant protein is retained in the endoplasmic reticulum (ER) and is not trafficked to the cell surface (misprocessed mutant). Corrector molecules such as corr-2b or corr-4a are small molecules that increase the amount of functional CFTR at the cell surface. Correctors may function by stabilizing CFTR at the cell surface or by promoting folding in the ER. To test if correctors promoted folding of CFTR in the ER, we constructed double cysteine CFTR mutants that would be retained in the ER and only undergo cross-linking when the protein folds into a native structure. The mature but not the immature forms of mutants M348C(... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594764 Tue, 25 Mar 2008 04:00:00 +0100 1594764 http://www.medworm.com/index.php?rid=1594768&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20070801 This study focuses on the 65 kDa glycoprotein Tr65 from the trout Onchorhynchus mykiss. Enzymatic digestion of Tr65 yielded a fragment pattern with strong homology to that of trout type II cytokeratin. Sequence analysis of the cDNA clone obtained by PCR confirmed this homology. We thus constructed a plasmid to overproduce recombinant Tr65. We extracted and purified this recombinant Tr65, using it for multi-channel and single-channel experiments in azolectin bilayers. Our results with recombinant Tr65 confirmed the pore-forming properties already shown with native antibacterial Tr65. These findings offer new insight into the function of keratin proteins present in various mucosal surfaces of animals and human beings. (Source: BJ Disease) BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594768 Thu, 20 Mar 2008 04:00:00 +0100 1594768 http://www.medworm.com/index.php?rid=1594767&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20071043 In this study we investigated the effect of lysosomal storage on bis(monoacylglycero)phosphate in cultured fibroblasts from patients with eight different lysosomal storage disorders and plasma samples from patients with one of 20 lysosomal storage disorders. Using electrospray ionization tandem mass spectrometry we were able to demonstrate either elevations or alterations in the individual species of bis(monoacylglycero)phosphate, but not of phosphatidylglycerol, in cultured fibroblasts. All affected cell lines, with the exception of Fabry disease, showed a loss of polyunsaturated bis(monoacylglycero)phosphate species relative to monounsaturated species and this correlated to literature reports of lysosomal dysfunction leading to elevations of glyc... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594767 Thu, 20 Mar 2008 04:00:00 +0100 1594767 http://www.medworm.com/index.php?rid=1594766&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20071657 It is known that p53 alterations are commonly found in tumors cells. Another marker of tumorigenesis is a Focal Adhesion Kinase (FAK), a non-receptor kinase that is overexpressed in many types of tumors. In our previous report we determined that the N-terminal domain of FAK physically interacted with the N-terminal domain of p53. In the present study by phage display, site-directed mutagenesis, pull-down and immunoprecipitation assays we localized the site of FAK binding to a 7 amino-acid region (65-71 a.a.) in the N-terminal proline-rich domain of human p53. Mutation of the binding site in p53 reversed the suppressive effect of FAK on p53-mediated transactivation of p21, BAX and Mdm-2 promoters. In addition, to functionally test this p53 site, we conjugated p53 peptides (wild type, contai... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594766 Thu, 20 Mar 2008 04:00:00 +0100 1594766 http://www.medworm.com/index.php?rid=1594765&cid=s_37616_60_f&fid=37616&url=http%3A%2F%2Fwww.biochemj.org%2Fbj%2Fimps%2Frefer.htm%3FMSID%3DBJ20071643 Chronic oxidative stress has been associated with genomic instability following exposure to ionizing radiation (IR). However, data showing direct causal linkages between specific reactive oxygen species and the IR-induced mutator phenotype are lacking. The current study demonstrates that IR-induced genomically unstable cells (characterized by chromosomal instability and increased mutation and gene amplification frequencies) show a 3-fold increase in steady-state levels of hydrogen peroxide, but not superoxide. Furthermore, stable clones isolated from parallel studies showed significant increases in catalase and glutathione-peroxidase activity. Treatment of unstable cells with polyethyleneglycol-conjugated catalase [PEG-CAT] reduced the mutation frequency and mutation rate in a dose depende... BJ Disease journals http://www.medworm.com/rss/comments.php?id=1594765 Thu, 20 Mar 2008 04:00:00 +0100 1594765